Anti-microbial compounds of 2&#39;, 4&#39; substituted anilides of substituted nitrosalicylic acid for quadruped animals

ABSTRACT

A class of organic compounds is disclosed comprising substituted anilides of 3-tert. butyl-6-methyl-5-nitro-salicylic acid, characterized in that the anilide portion is substituted only in the 4&#39; position or only in the 2&#39;, 4&#39; positions from a relatively small number of monovalent substituents. Such compounds are useful in the control of microorganisms and especially as anthelmintics, that is, therapeutic agents for destroying parasitic life, such as intestinal worms. Certain of the compounds are especially effective against Nematodes such as Haemonchus and Trematodes such as Fasciola.

CROSS-REFERENCE TO RELATED APPLICATIONS

This application is a continuation-in-part of an application filed Aug.1, 1972, Ser. No. 277,076, now U.S. Pat. No. 3,839,443, which is acontinuation-in-part of an application filed Oct. 4, 1971, Ser. No.186,514, now abandoned. Other related U.S. patents include U.S. Pat.Nos. 3,801,637 and 3,888,890.

BACKGROUND OF THE INVENTION

The need for combating the growth of undesirable microorganisms,bacteria, insects, and the like is a continuing and increasing one. Manyorganic compounds have been suggested as a deterrent to such growth oras an effective destroyer of the undesired life.

As compared to a mere killing or destroying of microorganisms, etc., aquite different situation prevails when one is concerned with killingonly one of two cohabiting classes of living things without harming theother. A common example of this is undesirable parasitic infestation ona desirable, living, warm-blooded, animal.

More specifically, various types of worm parasites are found in mammalsof commercial importance to man. The most important are the parasites oflivestock, especially of ruminants, such as sheep, goats, and cattle.However, other ruminants are similarly affected such as oxen, deer,water buffalo, etc. The more significant parasites are the nematodes(worms) of the alimentary tracts and the trematodes (flukes) whichinfect the liver. The alimentary tract nematodes are principallyimportant insofar as they raduce the growth of the host animals andrender less efficient the consumption of feed by the animals. Thetrematodes directly affect a vital organ and can cause severe illnessand death in the host animal.

Obviously, a treating agent which not only kills parasites, but alsokills the host animal is of no utility. Conversely, a treating agentthat is harmless on the host animal but only slightly retards the growthof parasitic life is of little real value. What is needed is a treatingagent that not only kills or expels the parasites, but which is harmlessto the host; or for which the host has a large margin of tolerance, thatis, an agent of which the host can take massive dosages with little orno harm. Moreover, although both types of mentioned infestations, thenematodes and the trematodes, occur commonly and naturally in the sametypes of livestock, presently known medications normally used forcontrol on one of these infections is generally ineffective for thecontrol of the other.

The matter of tolerance of a host animal for a therapeutic agent, suchas an anthelmintic, cannot be overemphasized, especially when the animalmust be administered to from a group or herd of animals in anunavoidably somewhat imprecise manner. In the treatment of large groupsthere is serious risk that some animals may be inadvertantly treatedmore than once, and thus subjected to double or triple dosage; that someanimals will be overdosed because of errors in estimating theirindividual weights; that some animals will, by virtue of individualgenetic variation, and variable state of health or ability, have lesstolerance than the average animal for any medication, It is thereforeclearly desirable that the typical animal be able to tolerate withoutserious harm, as large a multiple as possible of the minimum dosageregarded a likely to be effective as an anthelmintic.

The difference in activity between an effective amount of ananthelmintic on parasites and on a host animal can be quantitativelyexpressed as a Therapeutic Index. This index is defined as the maximumdose at which no toxic symptoms in the host animal are observed, dividedby the minimum dose at which the anthelmintic is therapeuticallyeffective. In general, an anthelmintic is considered to betherapeutically effective against a given parasite when it kills orexpells from the host at least 80% and preferably close to 100% of theviable forms of that parasite.

SUMMARY OF THE INVENTION

It has now been discovered that a class of compounds comprisingsubstituted anilides of 3-tert.-butyl-6-methyl-5-nitrosalicylic acidhave particular utility as a control for various types of microorganismsand also make effective anthelmintics against both nematodes andtrematodes when substituted only in the 4' position on the anilidemoiety, or when substituted only in the 2' and 4' positions, and noother positions. Additionally, the substituents themselves must beselected from a rather limited class. Still further, when thesesubstituents are reduced to an even more limited class, the resultingcompounds make superior anthelmintics. The latter compounds are not onlyhighly efficient in killing a wide spectrum of parasites but can betolerated without harm by host animals in relatively large amounts.

A compound of the present invention has the general formula: ##SPC1##

in which R₁ is H, chloro, bromo, iodo, CF₃, or alkyl of 1 to 4 carbonatoms, such as methyl or ethyl, and R₂ is chloro, bromo, iodo or CF₃.

The compounds of Formula I in which R₁ is methyl or CF₃, and R₂ ischloro, bromo, iodo or CF₃ are preferred. The compounds in which R₁ isCF₃ and R₂ is chloro or bromo are most advantageous.

DESCRIPTION OF THE PREFERRED EMBODIMENTS

Here and in the claims the following number system of salicylanilides isused: ##SPC2##

The salicylic acid portion of all compounds of the present inventionhave the same molecular configuration to afford the most usefulcompounds, namely, hydroxyl at the 2 position as indicated, tert. butylat the 3 position, nitro at the 5 position, and methyl at the 6position. The anilide portion of all compounds of the present inventionare substituted in only one or only two specific positions and at noother positions, namely, at the 4' position only or at the 2' and 4'positions only.

Additionally, the substituents at the 2',4' positions must be selectedfrom a relatively small group of monovalent substituents. As indicated,referring to the Formula I of the preceding section, R₁ must be H,chloro, bromo, iodo, CF₃, or alkyl of 1 to 4 carbon atoms, and R₂ mustbe chloro, bromo, iodo or CF₃. Anthelmintics of superior properties andperformance are obtained when R₁ is restricted to CF₃ and R₂ isrestricted to chloro or bromo.

Preparation of compounds of the present invention are described in thecited parent application, Ser. No. 186,514. In general, 3-tert.butyl-6-methylsalicylic acid may be nitrated in toluene solution withdropwise addition of nitric acid for about an hour at about 65°C. toabout 70°C. Upon cooling, 3-tert. butyl-6-methyl-5-nitrosalicylic acidprecipitates and may be recovered by filtration.

The anilide of the described salicylic acid may then be formed byreacting the acid with a phenylamine, the phenyl nucleus having thedesired substituent or substituents. This reaction may be carried out intoluene solution at 110°C. under reflux conditions for four to six hoursin the presence of a condensing agent such as POCl₃ or PCl₃. Uponcooling, a precipitate of a substituted anilide of3-tert.butyl-6-methyl-5-nitrosalicylic acid forms which can be recoveredby filtration, washing, and drying.

The new method of controlling helminths or parasitic worms by thisinvention comprises administration to a host animal such as a ruminantanimal in need of such treatment orally alone, combined with apharmaceutical or a feed carrier, an effective anthelmintic butnon-toxic quantity of a compound of Formula I. In certain cases such asextragastrointestinal infestation of flukes as in sheep, the compoundsmay be administered parenterally, that is, in a sterile micronizedsuspension or solution.

The administration is in quantities nontoxic but effective either forcurative or prophylactic purposes and has broad range of activity ongastrointestinal parasites of warm-blooded animals especially sheep andcattle. The helminths most effectively treated with the new compoundsare the Trematodes, Cestodes or, and especially, Nematodes. Activityagainst flukes such as Fasciola gigantica or Fasciola hepatica is alsoparticularly pronounced as noted above. More specific parasiticinfestations in which this invention may be applied are found in theMerck Veterinary Manual, Third Edition, pages 699-806, as are generalmethods of control of internal parasites; see also U.K. 1,183,641.Generally effective but nontoxic dose ranges of the active ingredientsare selected from about 0.25 mg./kg. of body weight up to about 50mg./kg. and preferably about 0.5 to 10 mg./kg. Effective dosages insheep without significant side effects have been found to be about 0.25to 50 mg./kg., with the monosubstituted compounds at R₂ (R₁ beinghydrogen) being less toxic and less active within this range. Thedisubstituted compounds are the preferred compounds and, as comparedwith the monosubstituted compounds, are much more active and somewhatmore poorly tolerated, although with a better therapeutic ratio. Mostusually the dosage unit compositions are administered from 1 to 5 timesdaily, preferably for convenience one treatment is used to clear theinfection. The dose range for the preferred compounds, in which R₁ istrifluoromethyl and R₂ is chloro or homo, is from about 0.25 to 25mg./kg. of body weight and preferably about 0.5 to 10 mg./kg. Theadvantageous bromo congener is used most generally at about 3 mg./1 kg.,usually as a drench, paste, bolus or top dressing.

Veterinary or compositions containing sufficient quantities of thecompounds of Formula I to reach the dose levels mentioned above areprepared as known to the art by preparing tablets, capsules, boluses,liquid suspensions, powders, drenches or solutions for injection inpackaged form. Alternatively, especially for prophylaxis, premix or feedcompositions containing effective but nontoxic quantities of the activesalicylanilide are used. For these purposes particulate carriers, inertpowders or, especially, feed carriers such as soybean meal, corn oil,vermiculite, diatomaceous earth, barley or wheat are used. In dosageunit or premix feed compositions the compound can comprise from about 5%to 75% by weight of the final composition as is convenient for thefarmer or veterinarian. As an example, a 5% salicylanilidevermiculite orsoybean meal premix can be used which will be uniformly mixed with theanimal foodstuff. Alternatively, a lick or pasture block can be used forfield animals.

To show the relative effect of compounds of the present inventiondiffering in the R₁ and R₂ substituents of Formula 1, as well as to showthe relative effect of those compounds with respect to still otherrelated compounds which are not of the present invention, that is, inwhich R₁ and R₂ are monovalent substituents other than those claimed, aseries of tests were carried out on living animals as follows.

Two groups of sheep were used in the tests. One group was used as anon-medicated control, while the other group was treated with thecompound under investigation. Helminth-free young lambs were infectedwith about 10,000 filariform larvae of Haemonchus contortus. In threeweeks when the infection became apparent, egg counts were made todetermine the density of the worm burden.

The egg count was carried out using the following modified Stoll method.Three grams of sheep feces were taken rectally from each animal andplaced in a Stoll flask. To each flask, tap water was added to the topline. The fecal pellets were too hard to break down, the flasks wereclosed with rubber stoppers and kept in the refrigerator for severalhours. The flasks were thoroughly shaken and a 1 ml. sample was drawnfrom the middle using a wide mouth pipette. This 1 ml. sample wastransferred to an 8 ml. plastic tube, to which enough saturated sodiumchloride solution was added to reach the rim. The tubes were thencentrifuged for 10 minutes at 1500 r.p.m. Then sodium chloride solutionwas added with a medicine dropper forming a concave miniscus. A coverglass was carefully added and the tubes were kept in a refrigerator forten to twenty minutes. During that period, the nematode ova flowedupward and adhered to the bottom of the cover glass slip. At the end ofthe 20-minute period, the cover slip was carefully detached from theminiscus and transferred to a microscopic slide. The ova were thencounted under a microscope equipped with 10X ocular and 10X objectivelens. Each ovum under the microscope represented 20 ova in the sample.

Animals with sufficiently high egg counts were taken out of the pool andhoused in experimental pens. Since the salicylanilides of the presentinvention are water insoluble, the compounds were first ground using amortar and pestle and then were placed in a carboxymethyl cellulosesolution. The particles were further micronized ultrasonically. Thecompound was then administered to two or three animals by Oesophagealgavage. Control, untreated animals, received only the carboxymethylcellulose solution in a similar manner. On the sixth and seventh dayafter treatment, fecal pellets were taken from the rectum of the controlas well as the treated animals and egg counts were made.

Test for activity against liver flukes were conducted as follows:

Eggs of Fasciola hepatica were collected from the bile of donor sheep.The eggs were embryonated and snails were infected (the genus Lymnaeaserves as intermediate host) to produce Metacercariae which are theinfective forms for sheep. Each sheep was infected with 250metacercariae intraruminally. When the infection became patent in ca. 80days, egg counts were carried out to determine the degree of wormburden. The liver was processed to recover F. hepatica in the untreatedand treated sheep.

The following examples illustrate the present invention and should notbe construed as imposing limitations upon the claims. Percentages are byweight percent unless otherwise indicated.

Although the present compounds are uniquely adapted for use asanthelmintics, they also find use in the control of various types ofmicroorganisms.

EXAMPLE 1

The following procedure for antibacterial evaluation was used. Testcompounds were dissolved in a suitable solvent (typically dimethylsulfoxide or acetone) and incorporated in a nutrient agar at variousconcentrations. The plates were then streaked with cultures of theappropriate bacteria. The inoculum contained about 8 × 10⁸ organisms permilliliter. After incubation for 48 hours at 37°C., the plates wereexamined for evidence of growth of the microorganisms. The minimumconcentration necessary for complete inhibition of growth was noted.

For testing against the fungus Trichophyton the same procedure was used,except that the test medium was Sabouraud agar, and the plates werecultured for 120 hours at 25°C. before evaluation for evidence ofgrowth. The inoculum added to the culture media contained about 5 × 10⁶spores in a water suspension containing 0.1% peptone.

Thus 4'-bromo-3-tert. butyl-6-methyl-5-nitrosalicylanilide was observedto have the following minimum inhibitory concentrations in agar platetests against bacteria.

    ______________________________________                                        Minimum Inhibitory Concentration (ppm)                                        ______________________________________                                        Staphylococcus aureaus    1                                                   Haemophilus gallinarum    8                                                   Escherichia coli         64                                                   Streptococcus foecalis   64                                                   Salmonella choleresuis   64                                                   Salmonella gallinarum    64                                                   ______________________________________                                    

The same compound was also found to be effective in reducing infestationon tomatoes by the fungi responsible for early blight (Alternariasolani) and late blight (Phytophthora infestans).

The compound, 3-tert. butyl-4'-chloro-6-methyl-5-nitrosalicylanilide, isinhibitory at 1 ppm to Trichophyton mentagrophytes, a fungus causingcertain mamalian skin disorders.

                  Example 2                                                       ______________________________________                                        Sheep Drench           Parts by Weight                                        ______________________________________                                        3-tert.Butyl-2', 4'-dichloro-6-methyl                                         5-nitrosalicylanilide  20                                                     Terra alba             75.5                                                   Tragacanth             3.0                                                    Sodium lauryl sulfate  1.5                                                    Water                                                                         ______________________________________                                    

The above solid components are mixed to give a water-dispersible powderto be used on concentrations of 5 grams of powder to 5 ml. of water. Thedrench is used orally as necessary and practical to controlgastrointestinal infections.

                  Example 3                                                       ______________________________________                                        Ruminant Bolus            Grams                                               ______________________________________                                        3-tert.Butyl-4'-chloro-2', 6-dimethyl                                         5-nitrosalicylanilide     0.5                                                 Calcium phosphate         4.0                                                 Maize starch              0.54                                                Talcum                    0.14                                                Gum arabic                0.15                                                Magnesium stearate        0.05                                                ______________________________________                                    

The phosphate and salicylanilide are mixed and screened, then granulatedusing one-half the starch. The screened and dried granules are mixedwith the remaining ingredients, blended thoroughly and compressed on abolus press.

Similarly, tablets can be prepared with reduced fillers.

EXAMPLE 4

The compounds of this invention also have utility against liver flukes.When administered an oral dosage of 5 milligrams per kilogram of bodyweight of sheep infected with liver flukes of the genus Fasciola,3-tert. butyl-2', 4'-dichloro-6-methyl-5-nitrosalicylanilide, completelydestroyed all flukes within three days. Against immature flukeinfestations in sheep similar activity was found at 15 mg./kg. The sameeffects against mature flukes were observed when 3-tert.butyl-4'-chloro-2', 6-dimethyl-5-nitrosalicylanilide was administered at5 milligrams per kilogram of body weight. The 4' bromo congener wasactive against mature flukes at 15 mg./kg. By contrast the isomericcompound 3-tert. butyl-3'-chloro 2', 6-dimethyl-5-nitrosalicylanilide isineffective against liver flukes even when the sheep receive 15milligrams per kilogram of body weight.

Other compounds of this invention which were tested and foundefficacious at 15 milligrams per kilogram of sheep body weight were:3-tert. butyl-6-methyl-5-nitro-4'-trifluoromethylsalicylanilide and3-tert. butyl-4'-chloro-6-methyl-5-nitrosalicylanililde.

In this respect, compounds of this invention differ from compoundscurrently in commercial use which are not effective against bothgastrointestinal worms and liver flukes. Since the simultaneousoccurrence of both forms of infections is common in commercial ruminanthusbandry, the value of a single form of medication to cure both typesof infection if self-evident.

EXAMPLES 5 THROUGH 13

The results of tests on sheep in a manner described, supra, usingcompounds of the present invention are given in Table 1, Examples 5through 13. The numbers in the column headed "Haemonchus (15 mg/kg)"refer to the percent of parasites destroyed or expelled when theindicated compound was introduced into the rumen of sheep at a dosage of15 milligrams per kilogram of body weight. The values under "Tolerance(mg/kg)" refer to the highest test dosage in the milligrams ofanthelmintic per kilogram of body weight at which the sheep developed notoxic symptoms. "Tol." is an abbreviation for "tolerated". The symbolsR₁ and R₂ refer to those of Formula I in these and other followingexamples.

                                      Table 1                                     __________________________________________________________________________    Substituants Haemonchus                                                                          Haemonchus                                                                          Haemonchus                                                                          Tolerance                                      Example                                                                            R.sub.1                                                                           R.sub.2                                                                           (2 mg/kg)                                                                           (5 mg/kg)                                                                           (15 mg/kg)                                                                          (mg/kg)                                        __________________________________________________________________________    5    Cl  Cl  14    100,99                                                                              100   >50,<100                                       6    CH.sub.3                                                                          Cl  67     95,99                                                                              100   >50,<100                                       7    H   Cl   0    65     81   >15,<50                                        8    H   Br  --    --     61   --                                             9    H   1   14    22     93   --                                             10   H   CF.sub.3                                                                          --    --    100   >15,<50                                        11   CF.sub.3                                                                          Br   99.8 --    --    Tol. at 20                                     12   CF.sub.3                                                                          Br  --     100  --    Tol. at 35                                     13   CF.sub.3                                                                          Cl  96    91    100   Tol. at 15                                     __________________________________________________________________________

It will be noted that the combination of substituents listed for R₁ andR₂ in Examples 5 and 6 are not only 100% effective in killing orexpelling Haemonchus, but dosages greater than 50 milligrams perkilogram of body weight and less than 100 milligrams per kilogram ofbody weight are tolerated by the sheep without harmful effects. Otherexamples also show highly satisfactory results.

In contrast to the good activity of the 2',4'-dichloro congener (Example5) 100% at 5 mg./kg. and tolerance between 50 and 100 mg./kg., andisomeric compound of the prior art namely the 2', 5'-dichloro (seeMonsanto British Pat. No. 1,252,087 published Nov. 3, 1971, firstcompound on page 5) has only 61% activity at 5 mg./kg. and shows toxiceffects at 50 mg./kg. It will be appreciated that only a 60% reductionin worm burden is not a practical or commercial objective for a newanthelmintic. The Monsanto patent also disclosed the salicylanilidesonly as larvicidal compounds against Lepidoptera or chewing insects. Itdiscloses no activity against internal parasites such as pin worms,round worms, flukes, etc. where a principal use of this invention lies.

EXAMPLES 14 THROUGH 22

When compounds of Formula I were used on sheep having substituents inthe 2' and 4' positions, other than those herein disclosed and claimed,the results were quite poor. Table II summarized the data for such othersubstituents.

                  Table II                                                        ______________________________________                                                                     Effectiveness in                                                              removal of Haemonchus                                                         at an intrarumenal                               Example   R.sub.1   R.sub.2  dosage of 15 mg/kg                               ______________________________________                                        14       CF.sub.3  H         0                                                15       F         H         0                                                16       Cl        H         33                                               17       Br        H         0                                                18       I         H         0                                                19       H         SO.sub.2 NH.sub.2                                                                       0                                                20       H         CO.sub.2 C.sub.2 H.sub.5                                                                18                                               21       CH.sub.3  CH.sub.3  13                                               22       OCH.sub.3 Cl        10                                               ______________________________________                                    

EXAMPLES 23 THROUGH 26

The importance of the 2',4' substitution on the anilide moiety ofFormula I is illustrated by the case of the dichloro substitution assummarized in Table III. It is evident from these data that the shift ofone or both of the chlorine atoms from the 2',4' configuration increasestoxicity and/or decreases efficacy. This has been mentioned brieflyhereinabove. As used herein, "toxic" refers to death of sheep resultingfrom the use of the indicated compound at the indicated dosage. The3',5' dichloro compound was not tested for tolerance since itseffectiveness was too low to be useful.

                                      Table III                                   __________________________________________________________________________    Position of                                                                   R.sub.1 and R.sub.2                                                                         Effectiveness (%)                                                                        Tolerance                                            Example                                                                            Substituents                                                                           5 mg/kg                                                                            15 mg/kg                                                                            50 mg/kg                                                                            100 mg/kg                                      __________________________________________________________________________    23   2',4'-dichloro                                                                         100  100   nontoxic                                                                            toxic                                          24   2',5'-dichloro                                                                         61   100   toxic --                                             25   3',4'-dichloro                                                                         20   100   toxic --                                             26   3',5'-dichloro                                                                         --    67   --    --                                             __________________________________________________________________________

EXAMPLES 27 THROUGH 30

Similarly, a shift of the substituents for the2'-methyl-4'-chloro-substituted anilide nolety of Formula I materiallyreduces efficacy and safety as shown by the following Table IV:

                                      Table IV                                    __________________________________________________________________________    Position of                                                                   R.sub.1 and R.sub.2                                                                            Effectiveness (%)                                                                        Tolerance                                         Example                                                                            Substituents                                                                              5 mg/kg                                                                            15 mg/kg                                                                            50 mg/kg                                                                            100 mg/kg                                   __________________________________________________________________________    27   2'-methyl-4'-chloro                                                                       95   100   non toxic                                                                           toxic                                       28   2'-methyl-3'-chloro                                                                       --   87    toxic --                                          29   2'-methyl-5'-chloro                                                                       --   0     --    --                                          30   2'-methyl-6'-chloro                                                                       --   0     --    --                                          __________________________________________________________________________

The tolerances of Examples 29 and 30 were not determined, since thecompounds were ineffective in combating the parasites in sheep.

EXAMPLES 31 THROUGH 34

For compounds of the present invention (Formula I) having hydrogen inthe 2' position of the anilide molety and chloro, homo, iodo ortrifluoromethyl in the 4' position, the effect of moving the4'-substituent is shown by the following Table V:

                                      Table V                                     __________________________________________________________________________    Effectiveness at 15 mg/kg against Haemonchus                                  Example                                                                            Substituent                                                                          4'-substitution                                                                         3'-substitution                                                                         2'-substitution                               __________________________________________________________________________    31   Cl     81        0         33                                            32   Br     61        --        0                                             33   I      93         00       0                                             34    CF.sub.3                                                                            100       2         0                                             __________________________________________________________________________

Thus, it is clear that a special and unpredicted superiority isassociated with substitution in the 4'-position.

EXAMPLES 35 THROUGH 39

The addition of a third substituent to a 2',4'-dichloro-or2'-methyl-4'-chloro-substituted anilide molety is also disadvantageousas shown by the following Table VI:

                  Table VI                                                        ______________________________________                                        Example Effectiveness at 15 mg/kg against Haemonchus                          ______________________________________                                        35      2',4'-dichloro    100%                                                36      2',4',5'-trichloro                                                                              Toxic                                               37      2',4',6'-trichloro                                                                              73%                                                 38      2'-methyl-4'-chloro                                                                             100%                                                39      2',5'-dimethyl-4'-chloro                                                                        0%                                                  ______________________________________                                    

EXAMPLES 40 THROUGH 49

The following Table VII provides physical characteristics includingcolor and melting points of some of the compounds of the presentinvention. These compounds are solids and were prepared by the processdescribed herein. The compounds are identified by their R₁ and R₂substituents in accordance with Formula I.

                  Table VII                                                       ______________________________________                                        Substituents                 Melting                                          Example                                                                              R.sub.1  R.sub.2  Color     Point °C                            ______________________________________                                        40     H        Br       Pale Yellow                                                                             136-138                                    41     H        I        Pale Yellow                                                                             143-145                                    42     H        Cl       Off White 150-152                                    43     H        CF.sub.3 Off White 175-176                                    44     H        I        Pale Yellow                                                                             143-145                                    45     CH.sub.3 Cl       White     167-168                                    46     Cl       Cl       Pale Yellow                                                                             150-152                                    47     H        F        Pale Yellow                                                                             205-206                                    48     CF.sub.3 Cl        --       152.5-153.5                                49     CF.sub.3 Br        --       161-163                                    ______________________________________                                    

EXAMPLES 50 THROUGH 60

These and the following examples illustrate the dual activity ofcompounds of the present invention against both Haemonchus and Fasciolahepatica as well as the relative inactivity of related compounds whichare not of the invention.

The data of the tables were generated using procedures previouslydescribed. The compounds tested are disubstituted and, except forExample 50 which is dibromo, are dichloro. Where the position for thesubstituents is other than 2' and 4', it is indicated. Under theindicated microorganisms, the values given in this and the followingtables, should be read as: percentage of microorganism killed/dose inmilligrams per kilogram of body weight. Under "Tolerance", the term"Tol." means "tolerated", "Tox." means "toxic", and "Sym." means"symptoms of toxic reaction", all at the dosage value which follows. Itwill be appreciated that there is no sharp toxicity end point indetermining toxicity to living sheep.

                  Table VIII                                                      ______________________________________                                        Substituents         Fasciola Tolerance                                       Example                                                                              R.sub.1 R.sub.2 Haemonchus                                                                            hepatica                                                                             (mg./kg.)                               ______________________________________                                        50     Br      Br      99.8/5  100/5  --                                      51     Cl      Cl      100/15  100/5   Sym./100                               52     Cl      Cl      100/5    99/2  Tol./50                                 53     Cl      Cl       14/2    44/1  --                                      54     Cl      Cl-5'   100/15  100/15 Tol./50                                 55     Cl      Cl-5'    61/5    0/5   --                                      56     Cl-3'   Cl      100/15  100/15 Tox./50                                 57     Cl-3'   Cl       20/5    95/5  Tox./15                                 58     Cl-3'   Cl       --      0/2   --                                      59     Cl-3'   Cl-5'   67,49/15                                                                              100/15 Tol./40                                 60     Cl-2'   Cl-3'    14/15   --    --                                      ______________________________________                                    

The 2',4' dichloro compound of the invention affords 100% dual activityat a 5 mg./kg. dose against both Haemonchus and Fasciola at atherapeutic ratio of at least 10:1. The 2',5' dichloro compound wouldnot be clinically effective against either parasite at 5 mg./kg., andextrapolation of the results indicate a clinically sufficient dose wouldbe achieved at a therapeutic ratio at a significantly lower level (about3:1 to 5:1). Against Fasciola, the 2',4' dichloro compound is 99%effective at doses 2.5 times less than that at which the 2',5' dichlorocompound was shown to have no activity.

EXAMPLES 61 THROUGH 73

The test data of these examples were obtained as before on compoundshaving methyl and halo substituents in which halo was chloro, homo, oriodo. The substitutions were at 2',4' unless otherwise noted.

                  Table IX                                                        ______________________________________                                        Substituents         Fasciola Tolerance                                       Example                                                                              R.sub.1 R.sub.2 Haemonchus                                                                            hepatica                                                                             (mg./kg.)                               ______________________________________                                        61     CH.sub.3                                                                              Cl      100/15  100/5   Tox./100                               62     CH.sub.3                                                                              Cl      99,95/5  0/2   Sym./50                                 63     CH.sub.3                                                                              Cl      67/2     --    Tol./25                                 64     CH.sub.3                                                                              Br      100/5   100/5  Tox./50                                 65     CH.sub.3                                                                              Br      38/2    100/5  Sym./50                                 66     CH.sub.3                                                                               I      88/5    100/15 Sym./50                                 67     CH.sub.3                                                                               I      13/2    100/5  --                                      68     CH.sub.3                                                                              Cl-3'   87/15    0/15  Tox./50                                 69     CH.sub.3                                                                              Cl-5'   0/15     --    --                                      70     CH.sub. 3                                                                             Cl-6'   0/15     --    --                                      71     Cl      CH.sub.3                                                                              62/15    --    --                                      72     Cl-3'   CH.sub.3                                                                              80/15    --    --                                      73     Cl-3'   CH.sub.3                                                                              0/5      --    --                                      ______________________________________                                    

Table IX demonstrates the much greater dual activity of the methyl 2',chloro 4' compound as well as its greater activity against Haemonchus.In many cases the methyl 2', chloro 4' isomer is very active at doseswell below those at which other isomers are inactive.

EXAMPLES 74 THROUGH 81

The data of these examples show in Table X the dual activity oftrifluoromethyl 2', and chloro 4' or bromo 4'.

                  Table X                                                         ______________________________________                                        Substituents         Fasciola Tolerance                                       Example                                                                              R.sub.1 R.sub.2 Haemonchus                                                                            hepatica                                                                             (mg./kg.)                               ______________________________________                                        74     CF.sub.3                                                                              Cl      100/15  100/2  Tox./25,50                              75     CF.sub.3                                                                              Cl       91/5   100/1  Tol./15                                 76     CF.sub.3                                                                              Cl       96/2    --     --                                     77     CF.sub.3                                                                              Cl       76/1    --     --                                     78     CF.sub.3                                                                              Br      100/5   100/5  Tol./35                                 79     CF.sub.3                                                                              Br      99.8/2  100/2  Tol./20                                 80     CF.sub.3                                                                              Br       --     100/1   --                                     81     CF.sub.3                                                                              Br       --     97/0.5  --                                     ______________________________________                                    

The trifluoromethyl 2', chloro or bromo 4' appear to be the most activecompounds of the invention, and of these the bromo 4' is preferred.

EXAMPLES 82 THROUGH 96

The data of these examples, shown in Table XI, are based onmonsubstitution in the position indicated on the anilide moiety.

                  Table XI                                                        ______________________________________                                        Example                                                                              Monosubstituent                                                                            Haemonchus                                                                              Fasciola hepatica                               ______________________________________                                        82            Cl-4'     81/15     100/15                                      83            Cl-4'     65/5      50/5                                        84            Cl-4'     0/2       0/2                                         85            Cl-3'     0/15      --                                          86            Cl-2'     33/15     --                                          87            CF.sub.3 -4'                                                                            100/15    100/15                                      88            CF.sub.3 -3'                                                                            2/15      --                                          89            CF.sub.3 -2'                                                                            0/15      --                                          90            Br-4'     61/15     100/15                                      91            Br-4'     --        0/5                                         92            Br-2'     0/15      --                                          93            I-4'      93/15      0/15                                       94            I-4'      22/5      --                                          95            I-3'      14/15     --                                          96            I-2'      0/15      --                                          ______________________________________                                    

The monsubstituted isomers of 4'-halo, consisting of chloro, bromo, andiodo, and of 4' trifluoromethyl of the present invention havesignificantly enhanced Haemonchus activity over the 2' and 3' isomers.

While the foregoing describes several embodiments of the presentinvention, it is understood that the invention may be practiced in stillother forms within the scope of the following claims.

I claim:
 1. The method for treating a parasite infested or parasiteexposed quadruped animal comprising administering orally to said animalan antiparasitically effective but nontoxic quantity of an activeingredient being a 3-tert.-butyl-6-methyl-5-nitrosalicylanilide of thegeneral formula: ##SPC3##in which R₁ is H, chloro, bromo, iodo, CF₃ oralkyl of 1 to 4 carbon atoms, and R₂ is chloro, bromo, iodo or CF₃. 2.The method claim 1 in which the administration is of a curative quantityto a parasite infested animal.
 3. The method of claim 1 in which theparasite is a Nematode or a Trematode and the quantity of activeingredient is chosen from about 0.25-50 mg./kg. based on the body weightof the host animal.
 4. The method of claim 1 in which the parasite is aHaemonchus or a Fasciola; R₁ is methyl, chloro, bromo, iodo or CF₃ andR₂ is chloro, bromo, iodo or CF₃ ; and the quantity of active ingredientis chosen from about 0.5-10 mg./kg. based on the body weight of the hostanimal which is either sheep or cattle.
 5. The method of claim 1 inwhich the parasite is a Fasciola; R₁ is CF₃ and R₂ is bromo; and thequantity of active ingredient is chosen from about 0.5-10 mg./kg. basedon the body weight of the host animal which is either sheep or cattle.6. The method of claim 5 in which the parasite is Fasciola hepatica andthe quantity of active ingredient is about 3 mg./kg. based on the bodyweight of the host animal.
 7. The method of claim 5 in which the activeingredient is in the form of a drench, paste, bolus or top dressing. 8.A veterinary composition for antiparasitic use in a host quadrupedanimal comprising an orally ingestable carrier and admixed therein as anactive ingredient an effective but nontoxic quantity of a3-tert.-butyl-6-methyl-5-nitrosalicylanilide of the formula: ##SPC4##inwhich R₁ is H, chloro, bromo, iodo, CF₃ or alkyl of 1 to 4 carbon atoms,and R₂ is chloro, bromo, iodo or CF₃.
 9. The veterinary composition ofclaim 8 in which said carrier contains a curative quantity of saidactive ingredient for a parasite infected host animal.
 10. Thecomposition of claim 8 in which the composition is for use against aNematode or a Trematode and the effective but nontoxic quantity ofactive ingredient is chosen from about 0.25-50 mg./kg. based on the bodyweight of the host animal.
 11. The composition of claim 8 in which theparasite is a Haemonchus or a Fasciola; R₁ is methyl, chloro, bromo,iodo or CF₃ and R₂ is chloro, bromo, iodo or CF₃ ; and the quantity ofactive ingredient is chosen from about 0.5-10 mg./kg. based on the bodyweight of the host animal which is either sheep or cattle.
 12. Thecomposition of claim 8 in which the parasite is a Fasciola; R₁ is CF₃and R₂ is bromo; and the quantity of active ingredient is chosen fromabout 0.5-10 mg./kg. based on the body weight of the host animal whichis either sheep or cattle.
 13. The composition of claim 12 in which theparasite is Fasciola hepatica and the quantity of active ingredient isabout 3 mg./kg. based on the body weight of the host animal.
 14. Thecomposition of claim 12 in which the active ingredient is in the form ofa drench, paste, bolus or top dressing.